Our major aim in these studies is to study the various phases of respiratory tract carcinogenesis at the molecular and cellular level using in vitro systems. We have previously developed an organ culture-cell culture system to demonstrate single and two-stage carcinogenesis in culture rat tracheal epithelial cells. We found that nontumorigenic epithelial cell lines could be established from explants exposed only to the powerful tumor promoter TPA (12-0-tetradecanoylphorbol-13-acetate). One of these cell lines was capable of metabolically activating benzo (a)pyrene (B(a)P) and its 7,8 dihydrodiol form to become malignantly transformed. The proximal carcinogenic metabolite of B(a)P, diolepoxide I, was more efficient in transforming the epithelial cell line than the intermediate or parent compound. Inoculations of the tranformed cell lines into immunosuppressed isogenic recipients produced differentiated carcinomas similar to those which occur in humans. These results indicate that tracheal epithelial cells can metabolically activate B(a)P, without the presence of other cell types, to become malignantly transformed. We are currently utilizing these epithelial systems to investigate mechanisms of chemical carcinogenesis and tumor promotion in airway epithelium.